There has been a significant research and development effort directed to the development of electrochemical sensors capable of detecting the presence and/or quantity of biologically significant analysis. Many, if not most of such analyte sensing devices are in the form of a test strip comprising a test fluid containment space pretreated with an analyte-dependent detection composition and electrodes for contact with test fluid delivered into the test fluid containment space. Electrical conductors extend from the electrodes to an area on the test strip for connection to a hand held or table mounted preprogrammed sensor reading device. Typically a biological fluid is delivered to the sample fluid containment area or volume and the sensor reading device is programmed to apply a predetermined potential to the electrodes after a predetermined period of time following delivery of the fluid sample to the sample containment space. Current flow is then measured responsive to said applied potential to provide an indication of the presence and/or concentration of the target analyte.
Some of such electrochemical sensors are constructed to prevent direct contact of the sample fluid with the electrode by covering the electrodes with a semipermeable membrane or gel matrix material, which is insoluble in the test medium and permeable at least to the analyte of interest when in contact with said test medium.
There is a continuing need for the development of commercially feasible, multi/continuous-use sensors for biologically significant analytes. In particular, there is need for development of biological sensors capable of being implanted or injected into a patient for periods ranging from several hours up to several weeks, months, or years and designed to provide accurate results without removal or recalibration to compensate for changes in diffusional properties of membrane components or for losses of enzyme activity and/or electron mediator elements. Such sensors would find application as components of artificial organs, for example an artificial pancreas, requiring continuous and/or regular monitoring of patient glucose levels. Such devices could also find use as reusable sensors for measuring analyte concentrations in bodily fluids in vitro, such as the analytical situations encountered by commercial labs performing analysis on patient fluid samples.
There are unique problems presented by the design and construction of biological sensors capable of repeated use in vitro and/or continuous use in vivo. Indeed, inherent in such functional requirements is the condition that the functional chemical component of the sensor be confined, i.e. not released from the sensor into the sample fluid during repeated and/or continuous use. The retention of the “active” chemical/electrochemical components of the biosensor can be accomplished by one of several techniques, alone or in combination. Thus the active components can be immobilized, for example, by covalent bonding to non-leachable components of the biosensor or by confining the biologically/electrically active components in a testing zone or volume by means of a membrane permeable at least by the analyte, but not by the contained, optionally covalently bound, enzymes, coenzymes, and/or electron mediators.
The implantable and/or reusable biosensors in accordance with the present invention are designed to retain the active sensor-dependent chemical components, typically in a hydrophilic matrix in an analyte retention volume. The active electrochemical species that cooperate in the sensor responsive to an applied potential to provide a current flow signal proportionate to the concentration of analyte diffuse into the retention volume can optionally be covalently bound to non-leachable components of the retention volume including, but not limited to, an electrode of an electrode system, a wall of the enclosure portion of the sensor for defining, at least in part, the retention volume, to microspheres or other microparticulate solids contained in the retention volume, to the retention volume contacting the side of a membrane, or to polymer components of the retention volume matrix. Alternatively the enzyme(s), the enzyme cofactor(s) and the electron mediator(s) can be selected to have a molecular weight sufficiently high to preclude any substantial diffusion of such components from the retention volume into the biological fluid being analyzed.
In one aspect of this invention the retention volume medium, alternatively denominated the “depletion volume medium” is in contact with the electrode system comprising an electrode capable of receiving electrons from or delivering electrons to the enzyme(s) via the electron mediator(s). Conductor elements extend from the electrode to a point on the device for allowing electrical communication of the electrode with a programmable controller. The controller can be programmed to apply a predetermined potential sequence to the electrode system including variable potential including either a mediator oxidizing potential or mediator reducing potential, variable pulse width and variable pulse intervals. The controller is also capable of sensing current flow responsive to applied potential(s) to the electrode system and comparing such data with control data previously obtained for said system to calculate and report analyte concentrations in the biological sample being analyzed and, optionally, to use such data to sense the performance status of the device and use such for modifying the then existing potential sequence protocol to optimize device function. Thus, for example, the sensor control can be modified periodically to adjust for differences in analyte diffusion efficiency across the membrane and/or changes in concentration of the active electron mediator and/or enzyme component(s) of the device without use of classical recalibration techniques.
In one embodiment of the invention the retention volume is defined or enclosed, at least in part, by an analyte-permeable membrane and the ratio of the retention volume to the surface area of the semipermeable membrane defining that volume, at least in part, is less than 2 mm, more preferably less than 1 mm. The low volume to surface area ratios are preferred in that they improve the rate of diffusional equilibrium between the fluid being tested and the retention volume medium, and thereby it works to minimize the refractory period (the recovery period) of the sensor.
In one preferred embodiment the enzyme component is selected so that it is substantially not capable of transferring electrons to or from any endogenous substance other than said analyte. Under such conditions the enzyme reaction responsible for providing a signal of analyte concentration cannot take place without a predetermined threshold potential being applied to the electrode system. The sensor can therefore be turned off to stop enzyme activity, optionally following a pulse of reducing potential to “deactivate” the mediator, and allow predictable concentration-gradient-based diffusion to work to rapidly “reset” the analyte concentration in the analyte detection/retention volume for the next programmed pulsed potential detection sequence.
In another embodiment of the present invention there is provided a method for monitoring analyte concentration using the sensor of this invention by contacting the sensor with the biological fluid being analyzed. Initially at predetermined intervals a potential is applied intermittently to the electrode system sufficient to oxidize the electron mediator in the retention volume, and the current flow through the electrode is sensed as a function of the duration of the applied potential. The applied mediator oxidizing potential is maintained at least for a period of time sufficient to determine the rate of change of current through the electrode as a function of duration of the applied potential. Values for the sensed current are correlated with values of current flow for known concentrations of the analyte. Alternatively the sensing protocol can comprise adjusting the potential to establish a predetermined current flow and thereafter sensing the rate of change of potential required to maintain said current flow for a predetermined time period.
In another embodiment the analyte concentration in a biological sample is measured as a function of the time dependent concentration of analyte in the retention volume following analyte depleting potential pulses. The rate of recovery concentration in the retention/depletion volume can be readily correlated with analyte concentration in the biological fluid contacting the sensor. The “diffusion status” of the membrane can be checked by a preprogrammed sequence from time-to-time during sensor use and numerical values associated with the sensed status can be used as input to modify the preprogrammed pulse sequence algorithms for subsequent sensor operation.
These and other features of the invention are described hereinbelow with reference to the drawings and the best mode known for carrying out the invention.